The gut microbiome is a key modulator of human health, metabolism, and immune function. Extensive study on the role of the microbiome has taken place since its significance was realized, with the majority of efforts characterizing its relationship to healthy and disease states. Unique stressors experienced by the Warfighter, such as sudden changes in diet and sporadic duration of fasted and fed states, have largely been unexplored. Here, we present our efforts to utilize in vitro fermentation as a modelling tool to simulate the gut microbial ecosystem and support Warfighter-centric gut research. In vitro fermentation affords greater flexibility and accessibility compared to animal or human clinical trials and can be utilized to inform in vivo studies. Recent efforts to develop a continuous culture model of the gut microbiome with separate bioreactors to represent each colonic domain will be presented. Use of an automated parallel bioreactor platform enabled real-time monitoring and control of numerous fermentation parameters including pH, anaerobicity, and fermentation volume. Inocula were prepared from fresh fecal donations of three individuals within 2 hrs of collection by processing in reduced phosphate buffer supplemented with glycerol and immediately storing at -20oC. Fermentation was performed utilizing 2% (w/v) fecal inocula in nutrient-rich media at 37oC under controlled anaerobic atmosphere. Colonic domains were simulated separately under batch conditions for 12-18 hours prior to being connected in sequence. The model then operated under continuous culture conditions, with earlier bioreactors in the sequence acting as feed reservoirs for later ones, for over five weeks. Aliquots were removed at discrete intervals to monitor phylogenetic population dynamics via denaturing gradient gel electrophoresis and 16S rRNA sequencing. Comprehensive characterization of phylogenetic changes is currently underway. During the final week of fermentation, a representative dietary input, purified cranberry proanthocyanidin, was introduced at multiple time-points to simulate intake during meals. Use of in vitro fermentation to model the human gut microbiome will enable broader investigation of the effect of Warfighter-relevant stressors on gut health and may act as a precursor to human or animal studies. Insight gleaned from this model, alone or in concert with in vivo studies, can inform nutritional strategies to restore and maintain Warfighter gut homeostasis.

Modeling the human gut microbiome through in vitro fermentation

254th National Meeting (2017)

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